Method and composition for the treatment of lipid and glucose metabolism disorders

ABSTRACT

Disclosed are methods for modifying or regulating at least one of glucose or lipid metabolism disorders which comprises administering to a human or vertebrate subject a D 1  dopamine agonist in conjunction with a dopamine D 2  agonist where the conjoined administration is effective to improve at least one of the following lipid and glucose metabolic indices: body weight, body fat, plasma insulin, plasma glucose and plasma lipid, and plasma lipoprotein. In preferred embodiments, the administration of the D 1  dopamine agonist and the D 2  dopamine agonist is conducted at a predetermined time.

[0001] This application claims priority under 35 U.S.C. §119 fromprovisional applications Ser. Nos. 60/017,377 and 60/019,336, thedisclosures of which are incorporated herein in their entirety.

FIELD OF THE INVENTION

[0002] This invention relates to novel, improved methods for modifyingor regulating in a subject (vertebrate animal or human) of at least oneof lipid and glucose metabolism.

BACKGROUND OF THE INVENTION Obesity and Lipid Metabolism Disorders—BodyFat Loss

[0003] In humans obesity can be defined as a body weight exceeding 20%of the desirable body weight for individuals of the same sex, height andframe (Salans, L. B., in Endocrinology & Metabolism, 2d Ed.,McGraw-Hill, New York 1987, pp. 1203-1244; see also, R. H. Williams,Textbook of Endocrinology, 1974, pp. 904-916). In other animals (or alsoin humans) obesity can be determined by body weight patterns correlatedwith prolactin profiles given that members of a species that are young,lean and “healthy” (i.e., free of any disorders, not just metabolicdisorders) have daily plasma prolactin level profiles that follow apattern characteristic of the species. This pattern is highlyreproducible with a small standard deviation. Members of a speciessuffering from at least one of lipid and metabolism disorders, however,have aberrant prolactin profiles that depart from the normal (or healthysubjects') pattern by at least 1 SEM in at least two spaced apart timepoints or by at least 2 SEM (standard error of the mean) in at least onetime point.

[0004] Obesity, or excess fat deposits, correlate with and may triggerthe onset of various lipid and/or glucose metabolism disorders, e.g.hypertension, Type II diabetes, atherosclerosis, etc.

[0005] Even in the absence of clinical obesity (according to the abovedefinition) the reduction of body fat stores (notably visceral fatstores) in man especially on a long-term or permanent basis would be ofsignificant benefit, both cosmetically and physiologically.

[0006] The reduction of body fat stores in domestic animals (as well aspets) especially on a long-term or permanent basis would also obviouslybe of considerable economic benefit to man, particularly since farmanimals supply a major portion of man's diet; and the animal fat may endup as de novo fat deposits in man.

[0007] Whereas controlled diet and exercise can produce modest resultsin the reduction of body fat deposits, prior to the cumulative work ofthe present inventors (including the prior co-pending patentapplications and issued U.S. patents referred to below), no trulyeffective or practical treatment had been found for controlling obesityor other lipid metabolism disorders.

[0008] Hyperlipoproteinemia is a condition in which the concentration ofone or more of cholesterol- or triglyceride-carrying lipoproteins (suchas chylomicrons, very low density lipoproteins or VLDL and low-densitylipoproteins or LDL) in plasma exceeds a normal limit. This upper limitis generally defined as the ninety-fifth percentile of a randompopulation. Elevated levels of these substances have also beenpositively correlated with atherosclerosis and the often resultingcardiac infarction, or “heart attack”, which accounts for approximatelyhalf of all deaths in the United States. Strong clinical evidence hasbeen presented which correlates a reduction in plasma lipoproteinconcentration with a reduced risk of atherosclerosis (Noma, A., et al.,Atherosclerosis 49:1, 1983; Illingworth, D. and Conner, W., inEndocrinology & Metabolism, McGraw-Hill, New York 1987). Thus, asignificant amount of research has been devoted to finding treatmentmethods which reduce levels of plasma cholesterol and triglycerides.High LDL and/or VLDL accompanied by high triglyceride levels in theblood constitute most important risk factors for atherosclerosis.Reduction of one or both of lipoproteins and triglycerides in the bloodwould reduce the risk of atherosclerosis and arrest or retard itsdevelopment.

[0009] Another subset of the plasma lipoproteins found in vertebratesare high density lipoproteins, or HDL. HDL serve to remove freecholesterol from the plasma. A high HDL concentration as a percentage oftotal plasma cholesterol has been associated with a reduced risk ofatherosclerosis and heart disease. Thus HDL are known in the lay pressas “good” cholesterol. Therefore, therapeutic strategies involveattempts both to reduce plasma LDL and VLDL content (that is, reducetotal plasma cholesterol), and to increase the HDL fraction of totalplasma cholesterol. Several lines of research indicate that simplyincreasing HDL is of benefit even in the absence of LDL or VLDLreduction: Bell, G. P. et al., Atherosclerosis 36:47-54, 1980; Fears,R., Biochem. Pharmacol. 33:219-228, 1984; Thompson, G., Br. Heart J.51:585-588,1989; Blackburn, H. N. E. J. M. 309:426-428, 1983.

[0010] Current therapies for hyperlipoproteinemias include a low fatdiet and elimination of aggravating factors such as sedentary lifestyle.If the hyperlipoproteinemia is secondary (i.e. incident to e.g. adeficiency of lipoprotein lipase or LDL receptor, various endocrinepathologies, alcoholism, renal disorders, hepatic disorders) thencontrol of the underlying disease is also central to treatment.Hyperlipoproteinemias are also treated with drugs, which usually alterthe levels of particular components of the total plasma cholesterol, aswell as reduce the total plasma lipid component. Among the most recentlyintroduced drugs to treat hyperlipoproteinemia is lovastatin (MEVACOR®)which selectively inhibits an enzyme involved in cholesterol production,3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase. This drugspecifically reduces total cholesterol and can cause a modest (5-10%)increase in HDL concentrations. However, benefits from these therapiesvary from subject to subject.

[0011] Moreover, use of the HMG-CoA enzyme inhibitor is sometimesaccompanied by side effects such as liver toxicity, renal myoglobinuria,renal shutdown, and lenticular opacity. The risk of such side effectsnecessitates close monitoring of the patients (e.g., liver function istested monthly).

[0012] Another drug prescribed against hyperlipoproteinemia isclofibrate. The effectiveness of clofibrate also varies from subject tosubject and its use is often accompanied by such side effects asnephrotic syndromes, myalgia, nausea and abdominal pain.

Diabetes and Glucose Metabolism Disorders

[0013] Diabetes, one of the most insidious of the major diseases, canstrike suddenly or lie undiagnosed for years while attacking the bloodvessels and nerves. Diabetics, as a group, are far more often afflictedwith blindness, heart disease, stroke, kidney disease, hearing loss,gangrene and impotence. One third of all visits to physicians areoccasioned by this disease and its complications, and diabetes and itscomplications are a leading cause of untimely death in the United Statesand in the Western world.

[0014] Diabetes adversely affects the way the body uses sugars andstarches which, during digestion, are converted into glucose. Insulin, ahormone produced by the pancreas, makes the glucose available to thebody's cells for energy. In muscle, adipose (fat) and connectivetissues, insulin facilitates the entry of glucose into the cells by anaction on the cell membranes. The ingested glucose is normally convertedin the liver to CO₂ and H₂O (50%); to glycogen (5%); and to fat(30-40%), the latter being stored in fat depots. Fatty acids from theadipose tissues are circulated, returned to the liver for re-synthesisof triacylglycerol and metabolized to ketone bodies for utilization bythe tissues. The fatty acids are also metabolized by other organs. Fatformation is a major pathway for carbohydrate utilization.

[0015] The net effect of insulin is to promote the storage and use ofcarbohydrates, protein and fat. Insulin deficiency is a common andserious pathologic condition in man. In insulin-dependent (IDDM or TypeI) diabetes the pancreas produces little or no insulin, and insulin mustbe injected daily for the survival of the diabetic. Innoninsulin-dependent (NIDDM or Type II) diabetes the pancreas retainsthe ability to produce insulin and in fact may produce higher thannormal amounts of insulin, but the amount of insulin is relativelyinsufficient, or less than fully effective, due to cellular resistanceto insulin.

[0016] In either form of diabetes there are widespread abnormalities. Inmost NIDDM subjects, the fundamental defects to which the abnormalitiescan be traced are (1) a reduced entry of glucose into various“peripheral” tissues and (2) an increased liberation of glucose into thecirculation from the liver. There is therefore an extracellular glucoseexcess and an intracellular glucose deficiency. There is also a decreasein the entry of amino acids into muscle and an increase in lipolysis.Hyperlipoproteinemia is also a complication of diabetes. The cumulativeeffect of these diabetes-associated abnormalities is severe blood vesseland nerve damage.

[0017] Other than the present invention and previous work by the presentinventors (discussed below), no effective treatment has been found forcontrolling either hyperinsulinemia or insulin resistance.Hyperinsulinemia is a higher-than-normal level of insulin in the blood.Insulin resistance can be defined as a state in which a normal amount ofinsulin produces a subnormal biologic response. In insulin-treatedpatients with diabetes, insulin resistance is considered to be presentwhenever the therapeutic dose of insulin exceeds the secretory rate ofinsulin in normal persons. Insulin resistance is also associated withhigher-than-normal levels of insulin i.e. hyperinsulinemia—when normalor elevated levels of blood glucose are present.

Previous Work in This Field

[0018] Studies by the present inventors and others have indicated thatthe naturally occurring annual cycle of body fat store level, pervasiveamong vertebrates in the wild, reflects the activities of an adjustablecentral metabolistat that is comprised of circadian hypothalamic neuralcomponents. Changes in the phase-relationships of circadian dopaminergicand serotonergic activities induce seasonal changes in metabolism andthese circadian activities can be adjusted by appropriately timedtreatments with hormones or neurotransmitter affecting drugs. In thisregard, bromocriptine, a sympatholytic dopamine D₂ agonist with α₂agonistic and α₁ antagonistic activities as well as serotonin inhibitingactivities has been demonstrated to reduce body fat store levels in avariety of animals including humans, without reducing food consumption,and also to reduce hyperinsulinemia, hyperlipidemia, and glucoseintolerance.

[0019] The present inventors and their co-workers have previously foundthat administration of either or both of (i) certain prolactin reducingdopamine (D₂) agonists such as bromocriptine and (ii)prolactin-increasing substances such as dopamine antagonists, such asmetoclopramide; and serotonin agonists and precursors, such as5-hydroxytryptophan, reduce body fat stores, obesity, plasmatriglycerides and cholesterol as well as hyperglycemia, hyperinsulinemiaand insulin resistance: U.S. Pat. Nos. 4,659,715; 4,749,709; 4,783,469;5,006,526.

[0020] It is preferred to administer the prolactin reducing substancesat a first predetermined time to effect a decrease in the circulatingprolactin levels of the subject to be treated during an interval withinthe subject's daily prolactin cycle or rhythm when circulating (blood)prolactin levels are low in young, healthy subjects of the same speciesthereby causing the prolactin rhythm of the treated to approach or toconform to the standard or healthy subjects' prolactin rhythm. It isalso preferred to administer the prolactin-increasing substances at asecond predetermined time to effect an increase in the circulatingprolactin levels of the subject to be treated during an interval withinthe subject's daily prolactin cycle or rhythm when circulating (blood)prolactin levels are high in young healthy subjects of the same species,thereby causing the prolactin rhythm of the treated subject to approach,or conform to, the standard or healthy subjects' prolactin rhythm. U.S.Pat. Nos. 5,468,755; 5,496,803; 5,344,832, 5,585,347 and U.S. patentapplication Ser. No. 08/456,952 and PCT applications US93/12701 andUS95/09061.

[0021] It is also known in the art that some of the effects ofbromocriptine are supported by endogenous dopamine. (Ergot Compounds andBrain Functions Neuropsychiatric Aspects: Advances in BiochemicalPsychopharmacology. M. Goldstein et al., Eds. (Raven Press, New York,1980) vol. 23). Specifically, it has been shown that locomotorstimulation and stereotyped behavior responses to bromocriptine areblocked by depletion of endogenous dopamine in rodents. However, if a D₁agonist is subsequently provided to dopamine depleted animals, theresponsiveness to bromocriptine is restored. Jackson, D. M. et al.,Psychopharmacology 94:321 (1988)). A similar dopaminergic D₂:D₁interaction has been demonstrated in dopaminergic inhibition of feedingbehavior. Although these studies confirm the importance of a D₂:D₁interaction in the activation of dopaminergic activities, the increasedlocomotor activity and decreased feeding response to D₂:D₁ agonists isacute and short lived, lasting for only a few hours. (Cooper, S. J. etal., in D ₁ :D ₂ Dopamine Receptor Interactions, J. Waddington, Ed.(Academic Press, London, 1993) pp. 203-234).

[0022] The previous work by third parties with D₁ and D₂ dopamineagonists in combination has not demonstrated any effects on lipid andglucose metabolism, and has not produced long-term responses ofdopaminergic activities. Significantly, the present inventors have nowfound that the conjoined administration of a D₁ agonist and a D₂dopamine agonist (or at least one of an adrenergic α₁ antagonist, anadrenergic α₂ agonist and a serotonergic inhibitor) result in anunexpected and surprising improvement in one or more of the metabolicindices related to lipid and glucose metabolism when compared to theimprovement (if any) provided by administration of a dopamine D₂ agonistsuch as bromocriptine administered alone.

OBJECTS OF THE INVENTION

[0023] It is one of the objects of this invention to provide additionalimproved methods for reducing in vertebrate subjects (including humans)in need of such treatment at least one of food consumption, body weight,body fat, plasma or blood glucose and blood insulin.

[0024] Another object of this invention is to provide methods forreducing at least one of insulin resistance (impaired glucosetolerance), hyperinsulinemia and hyperglycemia, and glycosylatedhemoglobin (including A1C), and abating Type II diabetes.

[0025] A further object is to provide methods for reducing or retardingor arresting atherosclerosis by reducing at least one ofhyperlipoproteinemia and elevated blood triglycerides.

[0026] It is another object of this invention to provide methods formodifying and regulating lipid and glucose metabolism in a mannerbeneficial to the subject.

[0027] It is still another object of the invention to provide methodsfor modifying and regulating lipid and glucose metabolism to provideeffective treatments for obesity.

SUMMARY OF THE INVENTION

[0028] It has now been found that at least one of the foregoing objectscan be accomplished by administering to a subject in need of suchtreatment a dopamine D₁ agonist in conjunction with one agent or agentcombination selected from the following:

[0029] (i) a dopamine D₂ agonist;

[0030] (ii) at least one of an adrenergic α₁ antagonist, an adrenergicα₂ agonist and a serotonergic inhibitor;

[0031] (iii) a dopamine D₂ agonist further conjoined with at least oneof an adrenergic α₁ antagonist, an adrenergic α₂ agonist and aserotonergic inhibitor.

[0032] Preferably, the foregoing agents in (i), (ii) or (iii) above(“conjoined agents”) are administered at a predetermined time i.e.within a restricted portion of a 24-hour period. Since the dopamine D₁agonist amplifies the effect of the other agent or agents, the D₁agonist is also preferably administered at about the same time.

[0033] The conjoined administration of a dopamine D₁ agonist with one(or more) of the other agents identified above results in substantiallyaugmented, and in fact often synergistic, effects in improvement of oneor more metabolic indices related to glucose or lipid metabolism, andthus an improved modification or regulation of at least one of glucoseand lipid metabolism.

[0034] Where a D₂ agonist is employed, it is preferably an ergotalkaloid, most preferably bromocriptine.

[0035] In another aspect, the present invention is directed toadministering to said subject:

[0036] (i) a D₂ agonist; and

[0037] (ii) at least one agent, not a D₂ agonist, selected from thegroup consisting of adrenergic α, antagonists, adrenergic α₂ agonistsand serotonergic inhibitors.

[0038] It has been found that such conjoined administration effects agreater improvement on one or more of the foregoing metabolic indicesthan administering of a D₂ agonist singly.

BRIEF DESCRIPTION OF THE FIGURES

[0039]FIG. 1 is a bar graph illustrating the weight loss (negative bars)or gain (positive bars) obtained in the experimental group administeredboth bromocriptine (BC) and SKF 38393 (SKF) compared to the animalsadministered SKF alone or BC alone or nothing (negative controls).

[0040]FIG. 2 is a graph of food intake (g/mouse/day) vs days oftreatment of experimental ob/ob mice with both bromocriptine and SKF(dark circles) or no drug (open circles) or control lean animals givenno drug (dark triangles).

[0041]FIGS. 3A and 3B are bar graphs measuring fat body mass measured asglycerol (in g/mouse) (FIG. 3A) or lean body mass (protein in g/mouse)(FIG. 3B) for ob/ob animals that received no drug (control) orbromocriptine alone (second bar from left) or SKF alone (third bar fromleft) or both BC and SKF (fourth bar). The asterisk indicates asignificant difference compared to the control bar.

[0042]FIGS. 4A and 4B are bar graphs of blood glucose (mg/dl) of ob/obanimals (FIG. 4A) or serum insulin (ng/ml) of ob/ob animals (FIG. 4B)administered no drug (control); left most bar); BC alone (second barfrom left); SKF alone (third bar from left) or both BC and SKF (fourthbar). The asterisks have the same significance as for FIG. 3A.

[0043]FIGS. 5A and 5B are bar graphs of serum triglyceride levels (TG)in ng/dl (FIG. 5A) or serum free fatty acid levels (FFA) in mmol/l (FIG.5B) for animals administered no drug (control; left most bar); BC alone(second bar from left); SKF alone (third bar from left) or both BC andSKF (fourth bar). The asterisks have the same significance as for FIG.3A.

[0044] FIGS. 6A-6C are bar graphs of blood glucose levels in mg/dl (FIG.6A) serum triglyceride levels in mg/dl (FIG. 6B) and serum FFA in mmol/l(FIG. 6C) for animals administered no drug (left bar) or both BC and SKF(right bar). The asterisks have the same significance as for FIG. 3A.The animals were sacrificed at 3 HALO the lipogenesis peak for mice.

[0045] FIGS. 7A-7C are bar graphs of liver enzyme activity (inmillimoles of fatty acid per mg protein per minute) for the enzymesinvolved in fatty acid synthesis in the liver: fatty acid synthetase(FIG. 7A), malic enzyme (FIG. 7B) or glucose-6-phosphatase (FIG. 7C)illustrating difference in said activities as between animalsadministered no drug (left bar) or both Bc and SKF (right bar). Theasterisks have the same significance as for FIG. 3A.

[0046]FIGS. 8A and 8B are bar graphs similar to those of FIGS. 7A-7C butfor the liver enzymes PEPCK (phosphoenol pyruvate carboxykinase) andglucose-6-phosphate dehydrogenase.

[0047] FIGS. 9A-9C are bar graphs similar to hose of FIGS. 7A-7C but forthe enzymes involved in fatty acid synthesis in adipose tissue: fattyacid synthetase (FIG. 9A) malic enzyme (FIG. 9B) and glucose-6-phosphatedehydrogenase (FIG. 9C).

[0048]FIGS. 10A and 10B are bar graphs of glucose transport (inamoles/cell/minute) (FIG. 10A) and glucose oxidation in CO₂ (inamoles/cell/minute) (FIG. 10B) measured for BC+SKF treated and “no drug”mice in the absence (white bars) and presence (dark bars) of insulin inisolated adipocytes.

[0049]FIG. 11 is a bar graph of lipolysis measured as glycerol release(pmoles/cell/minute) in isolated adipocytes for BC+SKF treated and “nodrug” mice.

[0050]FIGS. 12A is a graph of adipose lipogenesis measured as rate ofglycerol incorporation into lipids (mg/minute/gram of fat) as a functionof the sacrifice time for mice (in HALO) treated with BC+SKF (opencircles) or not treated (dark circles).

[0051]FIG. 12B is a bar graph of lipoprotein lipase (LPL) activity (inmmol of free fatty acid/10⁶ cells/hour for SKB+BC treated or “no drug”mice.

[0052]FIG. 13A and 13B are photomicrographs of adipocytes from BC+SKFtreated (FIG. 13B) and untreated (FIG. 13A) animals. The amount of lipidper cell (in μg lipid/cell) are given next to each Figure.

[0053] FIGS. 14A-14C are photomicrographs of arcuate nuclei of ob/obcontrol mice (FIG. 14A) ob/ob BC+SKF treated mice (FIG. 14B) and lean(57 BL/6J) controls (FIG. 14C) showing large amounts of neuropeptide Y(NPY) mRNA in the ob/ob controls and significantly reduced amounts ofNPY mRNA in the ob/ob treated mice.

[0054]FIG. 15 is a bar graph of NPY mRNA in the arcuate nucleus of ob/obmice treated with BC+SKF (middle bar) or untreated ob/ob mice (left bar)or untreated lean controls (right bar).

[0055]FIG. 16 is a plot of body weight v. day of treatment with a D₂agonist alone or with D₁ agonist alone or with a combination of D₁/D₂according to the invention. BC (10 mg/kg), BC plus SKF 38393, or vehicleinjection on body weight in C57BL/6J ob/ob mice during two weeks ofdaily treatment at 1 hour after light onset. An asterisk denotes asignificant difference in body weight change relative to all othertreatment groups (P<0.02).

DETAILED DESCRIPTION OF THE INVENTION

[0056] All literature and patents and patent applications cited hereinare incorporated by reference in their entirety. In case of a conflict,the present disclosure including its definitions shall control.

[0057] In one embodiment of the method of the present invention, a D₁dopamine agonist is administered in conjunction with a second agent,consisting of at least one of a D₂ agonist, an α₂ agonist, an α₁antagonist and a serotonergic inhibitor (or a D₂ agonist and at leastone of the remaining agents) preferably at a specific time of day to asubject in need of treatment.

[0058] As used herein and applied to administration of more than oneactive ingredient the terms “conjoined” or “in conjunction” mean thatthe subject being thus treated receives a first active agent and also atleast one other active agent, but not necessarily within the sameformulation or dosage form and not necessarily at the sameadministration time. For example, the D₁ agonist and D₂ agonist or theother agent(s) can be administered at the same time (in the same dosageform or in two or more divided dosage forms) or sequentially atdifferent times and in different dosage forms.

[0059] The D₁ dopamine agonist may be any one or more of thosesubstances known to those skilled in the art that are capable ofactivating or potentiating D₁ dopamine receptors. The D₁ agonists thatare suitable for use in the present invention include SKF38393,dihydrexidine, SKF 75670, SKF 82957, A77636, A68930, SKF 82526(fenoldopam), and racemic trans-10,11-dihydroxy5,6,6a,7,8,12b-hexahydro.

[0060] The D₂ agonists for use in the present invention can be any oneor more of those compounds known to those skilled in the art that arecapable of activating D₂ dopamine receptors. D₂ agonists suitable foruse in the present invention include LY-171555, bromocriptine methanesulfonate (+)-, 2,10,11-trihydroxyaporphine HBr, R(−)-, fisuridehydrogen maleate, 2-OH-NPA HCl, R(−)-, MDO-NPA HCl R(−),Propylnorpamorphine HCl R(−)-(NPA), and Quinperole HCl.

[0061] A preferred class of D₂ agonists includes ergot alkaloids such as2-bromo-alpha-ergocriptine (bromocriptine), 6-methyl 8 beta-carbobenzyloxy-aminoethyl-10-alpha-ergoline, 8-acylaminoergoline,6-methyl-8-alpha-(N-acyl)amino-9-ergoline, pergolide, lisuride,6-methyl-8-alpha-(N-phenyl-acetyl)amino-9-ergoline, ergocornine,9,10-dihydroergocornine, any D-2-halo-6-alkyl-8-substituted ergoline,and D-2-bromo-6-methyl-8-cyanomethylergoline. Of these bromocriptine ismost preferred.

[0062] Effective amounts of ergot alkaloid for humans and vertebrateswhen administered alone (not conjoined to a D₁ agonist) are typicallywithin the range of 5.0 ug/kg/day to 0.2 mg/kg/day.

[0063] In general, effective amounts of D₂ agonist for humans andvertebrates are within the range of 5ug/kg/day to 3.5 mg/kg/day.

[0064] The α₁ antagonists for use in the present invention can be anyone or more of those compounds known to those skilled in the art thatdirectly or indirectly block activation of α₁ adrenoceptors. The α₁antagonists suitable for use in the present invention includebromocriptine, benoxathin HCl, naftopidil 2 HCl, (i)-niguldipine HCl,S(+)-niguldipine HCl, prazosin HCl, doxazosin HCl, spiperone HCl,urapidil HCl, 5-methyl urapidil, WB4101 HCl.

[0065] Effective amounts of α₁ antagonist for humans and vertebrates aregenerally within the range of 0.02 to 0.3 mg/kg/day.

[0066] The α₂ agonists for use in the present invention can be any oneor more of those compounds known to those skilled in the art that arecapable of activating α₂ adrenoceptors. The α₂ agonists suitable for usein the present invention include bromocriptine, agmatine sulfate,p-aminoclonidine HCl, B-HT 920 diHCl, B-HT 933 diHCl, clonodine HCl,guanabenz acetate, p-iodoclonidine HCl, oxymetazoline HCl, UK 14,304,and xylazine HCl.

[0067] Effective amounts of α₂ agonist for humans and vertebrates aregenerally within the range of 1 ug/kg/day to 0.3 mg/kg/day.

[0068] The serotonergic inhibitors suitable for use in the presentinvention include bromocriptine.

[0069] Effective amounts of serotonergic inhibitors for humans andvertebrates are generally within the range of 5 ug/kg/day to 0.2mg/kg/day.

[0070] When two (or more) agents are administered in conjunction asdisclosed in the Summary of Invention the amount of one or another canbe lower than stated above, and even amounts that are subthreshold (whenan agent is used singly) can be employed.

[0071] The dopamine D₁ agonist and the dopamine D₂ agonist and/or otheragent conjoined with the D₁ agonist (or with the D₂ agonist) may beadministered to a subject preferably orally, or by subcutaneous,intravenous or intramuscular injection. Dermal delivery systems, e.g.,skin patches, as well as suppositories and other well-known systems foradministering pharmaceutical agents can also be employed. Sublingual,nasal and other transmucosal modes of administration are alsocontemplated. Accelerated release compositions, such as those disclosedin U.S. patent application Ser. No. 08/459,021, are preferred.

[0072] Each of the D₂ agonist, α₁ antagonist, α₂ agonist andserotonergic inhibitor are preferably administered at a predeterminedtime. The reason is that the effect of each of these agents on lipidand/or glucose metabolism is time-sensitive, as is explained in moredetail for D₂ agonists in in U.S. Pat. No. 5,585,347 and U.S. patentapplication Ser. No. 08/456,952, but applicable to the α₁ antagonists,α₂ agonists and serotonergic inhibitors. The preferred time ofadministration is within an interval that results in effective bloodlevels of the agent(s) at a time during which the standard prolactinlevels in healthy subjects of the species to be treated are low. Forexample in humans standard prolactin levels are low between the hours of9:00 and 22:00. Accordingly, the predetermined time of administration ofone or more of the foregoing agents is between the hours of 5:00 and13:00, preferably 7:00 and 12:00. Divided doses can be administered andthe schedule of administration can be varied to take into accountpharmocokinetic properties of each active agent. Details ofadministration are given in U.S. Pat. No. 5,585,347 and U.S. patentapplication Ser. No. 08/456,952 for bromocriptine, but also apply to theα₁ antagonists, α₂ agonists and serotonergic inhibitors employed in thepresent invention.

[0073] For mice the preferred time of administration of the active agentis within 1-hour after light onset. It is further preferred that theadministration take place when the subject is neither active norfeeding.

[0074] For other vertebrate animals the preferred time of administrationcan be ascertained by reference to the standard prolactin rhythm for thespecies of the animal to be treated. The standard prolactin curve can begenerated by measuring prolactin in young, healthy members of thespecies over a 24 hour period. See in U.S. Pat. No. 5,585,347 and U.S.patent application Ser. No. 08/456,952.

[0075] The administration of the D₁ agonist is also preferably timed,i.e. the D₁ agonist is also administered at a predetermined time.Because the D₁ agonist amplifies the effect of the conjoined agent, itis advantageous to administer the D₁ agonist at or about the time ofadministration of the conjoined agent(s), such that the activity periodof the D₁ agonist in the bloodstream of the treated subject overlaps (infact preferably overlaps as much as possible) with the activity periodof the conjoined agent. For convenience of administration and in orderto promote subject compliance, the D₁ agonist can be administered at thesame time as the conjoined agent(s).

[0076] The D₁ agonist may but need not be in the same formulation ordosage form (or form part of the same composition) as the conjoinedagent(s). If more than one conjoined agent is administered, theconjoined agents may but need not be in the same formulation or dosageform or form part of the same composition.

[0077] In treating vertebrates, generally, dosages of the D₁ agonist andconjoined agent(s) are typically administered over a period ranging fromabout 10 days to about 180 days, or longer. Some patients (e.g.,patients in particularly poor physical condition, or those of advancedage) may require a longer, or even continuous treatment. A treatmentduration exceeding six months or even continuous treatment may bedesirable even when not required.

[0078] At least one of body fat deposits, body weight, plasma or bloodglucose, circulating insulin, plasma triglycerides (TG), plasma freefatty acids (FFA) and food consumption of the subject will be reduced asthe result of the treatment. Disorders of lipid and glucose metabolismare thereby treated and subjects suffering from such pathologies ashyperphagia, obesity, insulin resistance (impaired glucose tolerance),hyperlipidemia, hyperinsulinemia, and hyperglycemia will exhibitimprovement in corresponding metabolic indices.

[0079] While appropriately timed administration of certain D₂ agonists(i.e., bromocriptine) alone will produce the effects described above tosome degree, these effects are amplified (potentiated) by the conjoinedadministration of the D₁ agonist agents described in the presentinvention. In other words, the synergistic effect of the conjoinedadministration of the D₁ agonist and the conjoined agent (i.e., a D₂agonist, and/or α₁ antagonist, and/or serotonergic inhibitor and/or α₂agonist) produces results that are superior to those experienced throughadministration of the same amount of a D₂ agonist alone. It should benoted that the present invention permits but does not require each agentto be administered in an amount over the threshold amount (in theabsence of a conjoined agent) to improve one or more metabolic indicesprecisely because of the augmented effect on these indices achieved byconjoined administration according to the present invention.

[0080] The benefits of the invention are not limited to modifying andregulating lipid and glucose metabolism. Other bodily functions, such asblood pressure, can be beneficially modified and regulated by timedadministration of a D₂ agonist (as monotherapy) in the dosage rangedisclosed above. For example, the D₂ agonist bromocriptine administeredat a dose within the range disclosed above (4.8 mg/day at 8:00 AM) hasbeen shown by the present inventor to decrease significantly thediastolic blood pressure of humans. Conjoined administration of adopamine D₁ agonist and

[0081] (i) a dopamine D₂ agonist;

[0082] (ii) at least one of an adrenergic α₁ antagonist, an adrenergicα₂ agonist, and a serotonergic inhibitor;

[0083] (iii) a dopamine D₂ agonist further conjoined with one or more ofthe members of (ii) above.

[0084] These and other features of the invention will be betterunderstood by reference to the experiments described in the examplesbelow.

EXAMPLE 1

[0085] Female ob/ob mice (40-70 g bw) were treated for two weeks witheither 1) bromocriptine (11 mg/kg) at light onset, 2) SKF38393 (20mg/kg) at light onset, 3) bromocriptine plus SKF38393 at light onset, or4) vehicle at light onset.

[0086] Bromocriptine or SKF38393 alone produced moderate reductions inhyperphagia, body weight gain, and obesity. However, bromocriptine plusSKF treatment produced significant reductions in hyperphagia (50-60%p<0.01) resulting in dramatic weight loss (21%, p<0.0001, compared tocontrols).

[0087] Body composition analysis of KOH/EtOH treated carcasses revealedno significant decrease of protein mass and a 22% (p<0.05) decrease ofadipose mass in bromocriptine plus SKF treated mice relative tocontrols. Also, bromocriptine plus SKF treatment decreased to a muchgreater extent than bromocriptine or SKF alone, plasma free fatty acid(FFA) (44%, p<0.001), triglyceride (TG) (50%, p<0.05), and glucose (57%,p<0.01). Insulin levels tended to decrease (by 50%; p<0.09) and totalcholesterol remained unchanged by combined drug therapy.

[0088] Larger (65-75 g) animals treated with bromocriptine plus SKF38393demonstrated an even more dramatic loss of body weight relative tocontrols (10±1 g in 10 days; p<0.01). Arcuate neuropeptide Y (NPY) mRNAlevels remain unchanged after bromocriptine plus SKF treatment comparedto controls.

[0089] C57BL/6J female obese mice of 40-45 g body weight were treated bydaily injections (at 1 HALO) of bromocryptine (BC at 10 mg/kg) and/orSKF38393 (SKF at 20 mg/kg). Animals were held on 12-hour dailyphotoperiods and fed ad libidum. Food consumption was monitored dailyand body weights monitored at days 0, 7 and 14 of the treatment.

[0090] Animals were sacrificed at 1 and/or 4 hours after light onset(“HALO”) (except as described for FIG. 12A) and blood, liver and adiposetissue were collected. The carcasses were digested in ethanolic KOH andanalyzed for protein and lipid content. Blood glucose was measured withan Accu-Chek Advantage glucose meter (Boehringer). Serum insulin wasmeasured with a radioimmunoassay kit (Linco Research) using rat insulinstandards. Total triglycerides and free fatty acids were measured withkits from Sigma Diagnostics, St. Louis, Mo. and Wako Chemicalsrespectively.

[0091] Enzymatic activity of fatty acid synthase, malic enzyme andglucose-6-phosphate dehydrogenase was measured in isolated cytosolicfraction by spectrophotometric methods. Phosphoenolpyruvatecarboxykinase (PEPCK) in liver cytosol was assayed by incorporation ofH¹⁴CO₃— into phosphoenolpyruvate. Glucose-6-phosphatase activity wasdetermined spectrophotometrically in isolated liver microsomes.

[0092] Adipocytes were isolated from perigonadal fat pads by collagenasedigestion and their size was determined by combining microscopesmeasurement of their diameter and lipid extraction of their lipidcontent. Glucose transport and glucose metabolism were measured byU-14C-glucose in the absence and presence of insulin and basal lipolysiswas assayed by measuring glycerol release using a ³²P-g-ATP.Neuropeptide Y (NPY) mRNA levels were measured in the arcuate nuclei ofthe mice using in situ hybridization.

[0093] In summary, bromocriptine (BC) plus SKF38393 (SKF) treatment ofC57 BL/6 ob/ob mice produces the following changes in metabolicphysiology:

[0094] (1) A 42% reduction in hyperphagia, reducing daily feeding levelsto less than or equal to lean (+/+) controls. (FIG. 2)

[0095] (2) A 3.67 g loss of body weight versus a 4.3 g body weight gainin obese controls. (FIG. 1)

[0096] (3) A 27% reduction of body fat mass (FIG. 3A) with no loss ofprotein (FIG. 3B) despite substantial reduction in food consumption.

[0097] (4) A 57 and 41 % reduction in hyperglycemia (FIG. 4A) andhyperinsulinemia (FIG. 4B) respectively.

[0098] (5) A 44 and 50% reduction serum FFA (FIG. 5B) and TG (FIG. 5A)concentration.

[0099] (6) A 27-78% reduction in lipogenesis enzymes within the liverand adipose (FIGS. 7A-7C, 8A-8B, 9A-9C).

[0100] (7) A 64% reduction in liver glucose-6-phosphatase and 80%increase in liver G6P dehydrogenase activities (FIG. 8B) as well assignificant reduction in fatty acid synthetase (FIG. 7A) and malicenzyme (FIG. 7B).

[0101] (8) A 42% reduction in basal lipolysis from isolated adipocytes(FIG. 11) of in vivo treated mice with no change in glucose transport(FIG. 10A) or oxidation (FIG. 10B) or GLUT4 expression (data not shown)as well as a significant reduction in adipocyte size (Compare FIGS. 13Band 13A).

[0102] (9) A 50% reduction in adipose tissue lipoprotein lipase (LPL)activity and a blocking of lipogenesis (FIGS. 12B and 12A respectively).

[0103] (10) The observed metabolic changes induced by BC+SKF areassociated with a 30% reduction in NPY mRNA level within the arcuatenuclei resulting in levels still two fold greater than in lean (+/+)counterparts (FIG. 15). A similar result can be qualitatively observedin FIGS. 14A-14C.

[0104] (11) The reduction in blood glucose, triglyceride and free fattyacid is more pronounced at 4 HALO (peak of lipolysis in the mouse). SeeFIG. 6 and compare FIG. 6A with FIG. 4A, FIG. 6B with FIG. 5A and FIG.6C with FIG. 5B.

[0105] Treatment of genetically obese C57 BL/6J mice with bromocriptine(D₂ agonist) plus SKF38393 (D₁ agonist) induced a reduction of bodyweight associated with a marked (42%) reduction of hyperphagia. Theresulting weight loss was attributed nearly exclusively to loss of fatwith protein mass remaining unchanged or even increased. Fat loss may beattributed to decreased caloric intake as well as decreased lipogenesisas both hepatic and adipose lipogenic enzyme activities were reduced bytreatment. The substantial reduction in caloric intake induced bytreatment was associated with a large reduction in circulating freefatty acids (FFA). That is, fat cell size (lipid content) decreasedappreciably while lipogenesis and lipid mobilization concurrentlydecreased. Apparently, the decreased mobilization is associated with aneven greater decrease in lipid accretion. Such a conclusion is supportedby the findings of decreased adipose LPL, and serum total and VLDL-TG(very low density lipoprotein/triglycerides).

[0106] The marked reduction in serum glucose induced by treatment isassociated with a strong reduction in hepatic glucose-6-phosphataseactivity and a somewhat less dramatic decrease in phosphoenolpyruvatecarboxykinase activity. Interestingly, the reduction in hepaticG-6-phosphatase activity and the simultaneous increase in G-6-Pdehydrogenase activity suggest specific metabolic channelling towardsglucose utilization in the liver rather than glucose release orproduction. Such alterations in liver metabolism facilitate increase ofhepatic HADPH, nucleic acid, and protein synthesis.

[0107] The foregoing findings may be applied to treatment of humanssuffering from obesity and other lipid disorders.

EXAMPLE 2

[0108] Different groups of 6-week old C57BL/6 ob/ob mice (lacking afunctional leptin protein) were treated with either bromocriptine (“BC”)(10 mg/kg BW), SKF38393 (“SKF”) (10 mg/kg BW), both drugs, or vehiclefor two weeks at 1 hour after light onset (HALO). Animals were held on12-hour daily photoperiods and allowed to feed ad libitum. Foodconsumption was monitored daily for 3 days before the initiation oftreatment throughout the 14-day treatment period. Animals weresacrificed between 1 and 3 HALO on the day following the final treatment(i.e., 24-26 hours after last injection) and plasma was collected forthe analyses of insulin, glucose, and lipids while the carcasses weresolubilized in ethanolic KOH and analyzed for protein and lipid content.Bromocriptine and SKF38393, individually, were ineffective in reducingbody weight gain where as SKF, but not BC, reduced food consumption(19%, P<0.01). However, the combined treatment of bromocriptine andSKF38393 (BC/SKF) decreased food consumption by 46% (from 4.8±0.2 to 2.6μg/day; P<0.001) and body weight by 15% (from a 3.2 g increase incontrols to a 4.3 g decrease; P<0.005) in 14 days of treatment (FIG.16). Relative to controls, in absolute terms, the lipid content of theBC/SKF treated animals was decreased by 40% (from 4.2±0.2 to 2.5±0.3 gglycerol/animal; P<0.0003) whereas the protein content increased 8%(from 3.7±0.08 to 4.0±0.08 g/animal; P<0.05). Therefore, relative tocontrol mice, the BC/SKF treated animals consumed less food but actuallyincreased protein mass while concurrently losing weight and fat. Thiseffect on body composition was observed by SKF (P<0.003) or BC (P<0.04)treatments alone, although to a lesser extent than by BC/SKF combination(P<0.05). Although BC alone and SKF alone significantly reduced plasmaglucose concentration (by 31%; P<0.02 and 43%; P<0.004, respectively),the BC/SKF combination reduced plasma glucose (by 60%; P<0.0004)substantially more than either drug alone (P<0.03) to values equivalentto those values reported for lean euglycemic C57BL/6 mice (+/+) (1).Plasma insulin level was equally reduced by BC and BC/SKF treatment(50%; P<0.04), but was not affected by SKF alone. BC/SKF, but neither BCnor SKF individually, reduced plasma triglyceride and free fatty acidlevels (by 36%; P<0.05 and 44%; P<0.007), (Table 1, below). These dataindicate that the interactive effects of BC and SKF effectively reducedhyperphagia, obesity, insulin resistance, hyperglycemia,hyperinsulinemia, and hyperlipidemia in the ob/ob mouse. TABLE 1 Effectsof BC (10 mg/kg), SKF (10 mg/kg), BC plus SKF, or vehicle injections at1 HALO on body weight, carcass composition, food consumption, and plasmaglucose, insulin, and lipid levels of ob/ob mice following two weeks oftreatment. Animals were sacrificed 24-26 hours following last treatment.Within parameters, values with similar superscripts denote a significantdifference between treatments (P < 0.05 to < 0.0001). Whole Body PlasmaPlasma Plasma Plasma Food Final Body Lipid-Glycerol Whole Body ProteinGlucose Insulin TG FFA Consumption Weight (g) (g) (% BW) (g) (% BW)(mg/dl) (ng/ml) (mg/dl) (uM) (g/day) CONTROL 54 ± 1¹ 4.2 ± 0.2¹ 7.9 ±0.3¹ 3.7 ± 0.1^(1,2,3) 6.5 ± 0.2¹ 380 ± 39^(1,2) 59 ± 12^(1,2) 313 ± 49¹825 ± 83¹ 48 ± 0.2^(1,4) BC 53 ± 0.7² 3.7 ± 0.1¹ 7.0 ± 0.2¹ 4.0 ± 0.1¹7.5 ± 0.1¹ 262 ± 25¹ 30 ± 4² 405 ± 101 818 ± 64² 4.5 ± 0.2^(2,5) SKF 52± 0.7² 3.1 ± 0.1¹ 6.0 ± 0.2¹ 4.1 ± 0.04² 7.9 ± 0.1¹ 218 ± 22² 50 ± 13234 ± 22¹ 671 ± 36² 3.9 ± 0.07^(3,4,5) BC/SKF 45 ± 2^(1,2,3) 2.5 ± 0.3¹5.4 ± 0.5¹ 4.0 ± 0.1³ 8.8 ± 0.0.4¹ 154 ± 15^(1,2) 30 ± 9¹ 199 ± 18 461 ±63^(1,2,3) 2.6 ± 0.2^(1,2,3)

EXAMPLE 3

[0109] The effects of BC/SKF treatment on circadian rhythms of keymetabolic enzyme activities, serum metabolites and hormones regulatingmetabolism were examined. Obese C57BL/6J mice were treated for 2 weeksat 1 hour after light onset with BC (10 mg/kg BW) and SKF (20 mg/kg BW)or vehicle. Mice were then sacrificed every 4 hours over a 24 hr periodfor the analyses of serum hormones and metabolites and hepatic enzymaticactivities. Serum glucose, free fatty acid (FFA) and hepaticglucose-6-phosphatase (G6Pase) activity were greatest during the lightperiod of the day showing that this time period is the daily peak forlipolysis and hepatic glucose production in mice. BC/SKF treatmentsignificantly reduced blood glucose (51%), FFA(56%) and G6Pase activity(38%) during this light period. Moreover, serum levels of the lipolyticand gluconeogenic hormones thyroxine and corticosterone were alsohighest during the light period and their levels were significantlyreduced by 51% and 53%, respectively by BC/SKF treatment. BC/SKFtreatment also decreased the daily peak in liver phosphoenol pyruvatecarboxykinase activity by 27% and increased the daily peak in liverglucose 6 phosphate dehydrogenase (by 32%) (potentiating glycolysis viaxylose-5-phosphate production). Levels of serum insulin and liver malicenzyme were greatest during the dark period (feeding time) of the dayillustrating increased lipogenesis during this time in mice. During thisdark period BC/SKF treatment reduced serum insulin significantly, i.e.,by 42%, and liver malic enzyme by 26%. BC/SKF treatment also decreasedliver fatty acid synthase activity by 30-50%, normalizing its circadianrhythm. Such effects demonstrate the involvement both of circadiansystems and BC/SKF influence on these systems in the regulation ofmetabolism.

EXAMPLE 4

[0110] The effect of in vivo BC/SKF treatment on glucose induced insulinrelease was studied in vitro. Obese (ob/ob) and lean (+/+) C57BL/6J micewere treated daily for 2 weeks with BC (10 mg/kg) plus SKF (20 mg/kg) orvehicle only. Mice were sacrificed 25 hours after the final treatmentand islets were isolated for static incubation with glucose. The BC/SKFtreatment of obese mice reduced blood glucose (173±14 mg/dl, P<0.01),plasma total glycerol 162±9 vs. 386±33 mg/dl, P<0.01), and plasma totalcholesterol (143±5 vs. 184±5 mg/dl, P<0.01) relative to obese controls.The plasma free fatty acid and insulin levels of treated mice were alsoreduced by 20-30% compared with that in obese controls. In control ob/obmice, the insulin release from isolated islets stimulated by 10 mMglucose was the same as that by 8 mM glucose (1.6±0.2 vs. 1.9±0.5ng/islet/h), while in BC/SKF treated ob/ob mice, 15 mM glucose induced asignificant increase of insulin release compared with 8 mM glucose(4.1±0.8 vs. 1.8±0.4 ng/islet/h, P<0.05). This enhancement is comparableto that observed in lean mice which exhibited a 2 fold increase ofinsulin release in response to 15 mM vs. 8 mM glucose. Similar BC/SKFtreatment of lean mice showed no effect on glucose-stimulated insulinrelease from isolated islets compared to lean controls. BC/SKF treatmentreversed impaired islet glucose sensing in ob/ob mice possibly due inpart to the improvement of hyperglycemia and hyperlipidemia by thistreatment.

[0111] Since hyperglycemia and hyperlipidemia may induce isletdesensitization to glucose, which is a common syndrome inobesity-associated NIDDM in humans, the above finding can be applied totherapy of NIDDM in humans.

EXAMPLE 5

[0112] Metabolic changes resulting from the D₁/D₂ agonist treatment wereevaluated in mice to determine if they were accompanied by decreases indensity of NPY immunoreactivity in discrete hypothalamic nuclei. Femaleob/ob mice (30-35 g) were treated daily at 1 h after light onset withSKF38393 (20 mg/kg) and bromocriptine (15 mg/kg) or vehicle. Lean mice(C57BL/6J; 18-21 g) treated with vehicle also served as controls.Following treatment for 12 days mice were sacrificed and their brainsprocessed for NPY immunoreactivity. The treatment (summarized in Table 2below) produced a significant decline in NPY levels in the SCN (38.5%P<0.01), the arcuate nucleus (41%; P<0.005) and the PVN (31.4% P<0.05)compared to obese controls. In addition, during the study body weightsincreased in obese controls (8.3±0.9 g) whereas it decreased in treatedanimals (−1.1±2 g) (P<0.0001). These results indicate that time ofday-dependent dopaminergic D₁/D₂ coactivation improves hyperphagia,hyperglycemia and obesity in the ob/ob mouse, in part, by reducingelevated levels of hypothalamic NPY to that of lean animals. TABLE 2Food Blood NPY density consumed glucose (arbitrary units) Type (g/day)(mg/dl) SCN Arcuate PVN Lean 3.1 +/− 133 +/− 5 39.8 +/− 3 54 +/− 4 49+/− 5 0.1 Obese 6.1 +/− 216 +/− 16 55.2 +/− 4 95 +/− 10 52 +/− 6 0.1Treated 4.3 +/− 136 +/− 9^(c)   34 +/− 4^(b) 56 +/− 8^(b) 36 +/− 3^(a)0.1^(c)

EXAMPLE 6

[0113] The influence of BC/SKF treatment on hepatic glucose metabolismwas examined. Female C57BL/6J obese (ob/ob) mice (BW=46±1 g) weretreated daily for 2 wks with BC (12.5 mg/kg) and SKF (20 mg/kg) orvehicle (n=8-12/ group) at 1 hr after light onset and then sacrificed at24-26 hrs following the final day of treatment and liver tissue removedand analyzed for glucose 6-phosphatase (G6Pase) and glucose 6 phosphatedephdrogenase (G6PDase) activities and hepatic xylose-5-phosphate (X5P)concentration. Serum glucose and insulin levels were also determined.BC/SKF treatment significantly (P<0.01) reduced serum glucose by 57%(from 435±21 to 185±8 mg/dl), serum insulin by 44% (from 25±2 to 14±3ng/ml), hepatic G6Pase activity by 67% (from 1.5±0.3 to 0.5± 0.07μmoles/min/mg), and increased hepatic G6PDase activity by 73% (from 11±1to 19±3 nmoles/min/mg), and X5P concentration by 73% (from 166±10 to287± 30 nmoles/g) relative to control. BC/SKF treatment resulted in agluconeogenic substrate being shuttled away from glucose to the pentosephosphate pathway by the simultaneous inhibition of glucose6-phosphatase (G6Pase) and stimulation of glucose-6-phosphatedephdrogenase (G6PDase) thereby respectively blocking hepatic glucoseproduction and shuttling glucose-6-phosphate towards production ofxylose-5-phosphate (X5P), a potent activator of glycolysis. This is thefirst study to identify the existence of such a biochemical shift inhepatic glucose metabolism and its regulation by dopaminergicactivation. Moreover, this dopaminergic regulated shift from hepaticgluconeogenesis towards potentiation of glycolysis could contribute tothe normalization of severe hyperglycemia in these animals and may havesignificance in both the development and treatment of NIDDM in humans.Available evidence suggests that BC/SKF is acting in part at theventromedial hypothalamus to produce these effects.

EXAMPLE 7

[0114] The combination of hyperglycemia and hypertriglyceridemia hasbeen implicated as a risk factor for cardiovascular disease in NIDDM.Dopaminergic D₁/D₂ receptor co-activation with SKF38393 (SKF), a D₁receptor agonist, plus bromocriptine (BC), a D₂ receptor agonist hasbeen shown to act synergistically to reduce obesity. Its effects onhyperglycemia, dyslipidemia, and plasma lipoprotein dynamics were testedin ob/ob mice. Obese C57BL/6J (ob/ob) mice (BW) 44.5±0.5 g) were treateddaily at light onset with vehicle (control) or SKF (20 mg/kg BW) plus BC(16 mg/kg BW) for 14 days. 25 to 28 hrs following the final treatment,animals were sacrificed and blood was collected for lipoproteinfractionation and analysis. Lipoprotein and serum triglyceride (TG),cholesterol (CH), phospholipid (PL), and serum glucose, insulin, andfree fatty acid (FFA) were measured. White adipose, skeletal muscle, andheart tissues were harvested for analysis of lipoprotein lipase (LPL)activities. A second group of similarly treated animals was utilized fordetermination of hepatic triacylglycerol synthesis by following³H-glycerol incorporation into liver triglyceride 30 mins after itsadministration in vivo. 14 days of SKF/BC treatment significantlyreduced blood glucose (390±17 to 168±6 mg/dl), serum TG (397±22 to 153±7mg/dl), CH (178±4 to 139±4 mg/dl), PL (380±7 to 263±11 mg/dl), and FFA(1.1±0.1 to 0.7±0.1 mmol/1) (P<0.01). Insulin was also reduced from 40±5to 28±4 ng/ml (P=0.058). Chylomicron-TG and VLDG-TG were reduced from228±2 to 45±6 mg/dl and 169±7 to 110±4 mg/dl, respectively (P<0.01).Hepatic triacylglycerol synthesis was reduced by 47% (P<0.01). LPLactivity was unchanged in skeletal and heart muscle tissues but wassharply reduced (67%) in adipose tissue (P<0.01). LDL cholesterol levelwas reduced by 31 % (P<0.01). These data indicate that SKF/BC normalizedhypertriglyceridemia via 1) decreasing Chylomicron-TG level and 2)decreasing VLDL-TG synthesis and secretion. The marked decrease inadipose LPL activity further supports the conclusion that serum VLDL-TGis reduced by decreased hepatic synthesis rather than increased removalfrom the circulation and may also contribute to the decreased serum FFAlevel. Moreover, the decreased serum FFA may contribute to the decreasedhyperglycemia.

EXAMPLE 8

[0115] A 2-week treatment with SKF38393 (SKF), a dopamine D₁ receptoragonist, and bromocriptine (BC), a dopamine D₂ receptor agonist actssynergistically to reduce body fat and hyperglycemia in ob/ob mice in afood consumption independent manner. The biochemical mechanismsresponsible for this effect were evaluated by measuring energyexpenditure and metabolic substrate utilization determined fromrespiratory quotient (RQ) of treated versus control mice. Circulatingfree fatty acid (FFA) levels represent the major rate limiting factorfor fat oxidation and increased FFA also potentiate hyperglycemia ininsulin resistant states. The influence of in vivo treatment withSKF38393 (SKF), a dopamine D₁ receptor agonist, and bromocriptine (BC),a dopamine D₂ receptor agonist, on serum FFA level and in vitrolipolysis in isolated adipocytes was tested. C57BL/6J obese (ob/ob)female mice were treated with vehicle (control) or SKF k(20 mg/kg BW)plus BC (10 mg/kg BW) for 14 days. BC/SKF treatment increased O₂consumption and CO₂ production by 143% and 90% respectively (P<0.0001).Moreover, RQ values were shifted by treatment from 1.55±0.35 to1.03±0.11 indicating a decrease in de novo glucose conversion to lipids(lipogenesis) and nearly exclusive utilization of glucose as an energysource (i.e., little fat oxidation). These findings are in accord withthe substantial drug-induced decrease in serum glucose level (489±25 to135±10 mg/dl, P<0.0001). These conclusions from the RQ data are furthersupported by a dramatic decrease in fat cell size (from 0.722±0.095 to0.352±0.03 μg of lipid/cell, P<0.02) and a marked reduction in serum FFAlevels (from 1.06±0.1 to 0.32±0.02 mM, P<0.001) and in vitroisoproterenol stimulated lipolysis (from 16.4±2.4 to 5±0.6 pmoles ofglycerol released/cell/20 min, P<0.005). Therefore, the dramaticincrease in O₂ and CQ production (and decreased fat cell size) cannot beexplained by increased fat mobilization and oxidation. These dataindicate that dopaminergic D₁-D₂ receptor coactivation shifts glucosemetabolism from lipogenesis to oxidation with a concurrent decrease offat mobilization and oxidation (thereby possibly improving insulinsensitivity). These findings have significance for the treatment ofobesity and hypertriglyceridemia associated with NIDDM.

EXAMPLE 9

[0116] The combined effectiveness of SKF38393 (SKF), a D₁ receptoragonist, and bromocriptine (BC), a D₂ receptor agonist, were examined intreating obesity and diabetes in ob/ob (mice lacking the gene for theleptin protein) and db/db (mice lacking the gene for the leptinreceptor) mice. Daily drug injections were administered to femaleC57BL/6J ob/ob and C57BL/KJ db/db mice 1 hr after light onset for 14days. Drug treated groups received BC (16 mg/kg) plus SKF (20 mg/kg),whereas pair fed groups (food adjusted to drug treated groups' intake)and control groups received the vehicle. Oxygen consumption was measuredin metabolic cages on day 11 or 12 of treatment. Plasma glucose, FFA,and insulin levels, were measured on day 14. In the ob/ob micestatistically significant results included: controls gained 6.9±1.3 g ofbody weight, while the treated mice lost 7.4± 0.4 g. The average dailyfood consumption of controls was 6±0.2 g versus 2.8±0.1 g of treated.Oxygen consumption for controls and treated was 1277±240 ml/kg/hr and1623±230, respectively. Plasma glucose levels were 471±42 mg/dl incontrols, and 164± 13 in treated. k FFa levels were 1.27±0.1 mM incontrols, and 0.37±0.05 in treated. Plasma insulin were 63.5±17 ng/ml incontrols, and 37.3±6.6 in treated. Similar statistically significantresults were observed in db/db mice: controls gained 6.6±0.4 g, of bodyweight versus 3.4±1.3 g in the treated. The average daily goodconsumption of controls was 10.7±2.8 g versus 5.9±0.5 g of the treated.Oxygen consumption for control and treated was 898±2150 ml/kg/hr and2322±283, respectively, Plasma glucose levels were 485±29 mg/dl incontrols, and 390±55 in the treated. FFA levels were 1.49±0.2 mM incontrols, and 0.45±0.04 in treated. Plasma from pairfed animals (in bothob/ob and db/db mice) indicate that the above drug-induced metabolicchanges are not primarily the consequence of decreased food consumption.These results strongly suggest that hyperphagia, hyperglycemia andhyperlipidemia in animals lacking either leptin (ob/ob) or a functionalleptin receptor (db/db) can be treated with the combined administrationof D₁ and D₂ receptor agonists.

EXAMPLE 10

[0117] Pharmacological intervention with bromocriptine improves glucoseand lipid metabolism in NIDDM animals and patients. The influence ofsuch treatment on pancreatic islet function was investigated. The effectof D₁/D₂ receptor agonists— bromocriptine/SKSF38393 (BC/SKF) on isletfunction in a mouse diabetic model was evaluated. Female db/db mice(30±1 g) were treated daily for 2 weeks at 1 hr after light onsetwith 1) BC (16 mg/kg) plus SKF (20 mg/kg), 2) vehicle only (controls),or 3) vehicle plus feed restriction to match the reduced foodconsumption of treated mice (pair fed). The BC/SKF treatment reducedblood glucose (347±28 vs. 606±31 mg/dl in controls, P<0.01) and plasmafree fatty acids (0.6±0.1 vs. 1.1 k±0.1 mM in controls, P<0.01) levels,and increased plasma insulin level by 3-fold compared with that incontrols (49±5 vs. 16±2 ng/ml, P<0.01). In pair fed mice there was amore modest (30%) reduction (P<0.01) of blood glucose but no change inplasma insulin and a 20% increase in plasma free fatty acids comparedwith control levels. The insulin release response of pancreatic isletsto secretagogue was tested in vitro. Insulin release from incubatedislets stimulated by glucose (8 and 15 mM), arginine (10 mM) andacetylcholine (10 μM) was each 34 fold greater in the treated groupcompared with that in controls (P<0.05). Contrariwise,secretagogue-induced insulin release from incubated islets of pair fedmice were similar to those in controls. Furthermore, similar BC/SKFtreatment had no effect in normal mice. Addition of BC/SKF directly tothe islet incubation buffer did not enhance insulin release from db/dbmouse islets. These results demonstrate the BC/SKF given in vivomarkedly enhance islet function in the db/db but not the normal mouse.This effect is not attributable to either a direct action on isletfunction or inhibition of feeding.

What is claimed is:
 1. A method for modifying or regulating at least oneof glucose or lipid metabolism disorders which comprises administeringto a human or vertebrate animal subject in need of such modification orregulation a D₁ dopamine agonist in conjunction with a dopamine D₂agonist wherein the conjoined administration is effective to improve atleast one of the following lipid and glucose metabolic indices: bodyweight, body fat, plasma insulin, plasma glucose and plasma lipid, andplasma lipoprotein.
 2. The method of claim 1 wherein the D₂ agonist isan ergot alkaloid is selected from the group consisting of2-bromo-alpha-ergocriptive, 6-methyl 8beta-carbobenzyloxyaminoethyl-10-alpha-ergoline, 8-acylaminoergoline,pergolide, lisuride, 6-methyl-8-alpha-(N-acyl) amino-9-ergoline,6-methyl-8-alpha-(N-phenyl-acetyl)amino-9-ergoline, ergocomine,9,10-dihydroergocomine, and D-2-halo-4-alkyl-8-substituted ergolines,D-2-bromo-6-methyl-8-cyanomethylergoline.
 3. The method of claim 1wherein the D₁ agonist comprises SKF38393.
 4. The method of claim 2wherein the ergot alkaloid comprises bromocriptine.
 5. The method ofclaim 1 wherein the D₂ agonist is administered at a predetermined timeof day.
 6. The method of claim 5 wherein the D₁ agonist is administeredat about the same time as the D₂ agonist.
 7. A method for modifying orregulating at least one of glucose or lipid metabolism disorders whichcomprises administering to a human or vertebrate animal subject in needof such modification or regulation a D₁ agonist in conjunction with atleast one member selected from the group consisting of an adrenergic α₁antagonist, an adrenergic α₂ agonist and a serotonergic inhibitorwherein the amount of the combination is sufficient to effect at leastone of the following lipid and glucose metabolic indices: body weight,body fat, circulating insulin, plasma or blood glucose, plasma lipid andplasma lipoprotein.
 8. The method of claim 7 wherein the D₁ agonist andthe member of said group is administered at a predetermined time of day.9. The method of claim 8 wherein the D₁ agonist is administered at aboutthe same time as member of said group.
 10. A method for modifying orregulating at least one of glucose or lipid metabolism disorders whichcomprises administering to a human or vertebrate animal subject in needof said modification or regulation a D₁ agonist in conjunction with atleast one of a α₂ agonist, a α₁ antagonist and a serotonergic inhibitor,and further in conjunction with a D₂ agonist sufficient to improve atleast one of the following lipid and glucose metabolic indices: bodyweight, body fat, plasma insulin, plasma glucose and plasma lipid.
 11. Atherapeutic agent combination for modifying or regulating at least oneof glucose and lipid metabolism disorders comprising: a first amount ofa dopamine D₁ agonist; and a second amount of at least one memberselected from the group consisting of: (i) a dopamine D₂ agonist; and(ii) at least one of an adrenergic α₁ antagonist, an adrenergic α₂agonist and a serotonergic inhibitor.
 12. A method for modifying orregulating at least one of glucose or lipid metabolism disorderscomprising: administering to a subject in need of said modification orregulation: (i) a D₂ agonist in a first amount; and (ii) at least one ofan adrenergic α₁ antagonist, an adrenergic α₂ agonist and a serotonergicinhibitor in a second amount; said first and second amounts beingsufficient to improve at least one of the following indices of glucoseor lipid metabolism: body weight, body fat, plasma insulin, plasmaglucose and plasma lipid.